In the fields of medicine and clinical chemistry, many studies and determinations of physiologically reactive species, e.g. cells, protein, enzymes, cofactors, nucleic acids, substrates, antigens, antibodies, etc. are carried out using "labels" which facilitate the detection or separation of the materials under observation at low concentration. In one such application, the diagnosis of pathological conditions and the detection of drugs or narcotics in humans and animals is often carried out using labeled materials in specific binding assays using competitive binding principles.
Whenever labels are used, sensitivity is of prime importance due to the generally low levels of biological species that are measured. Procedures carried out using radiometric labels generally do not have sufficient sensitivity for many low level analytes. In addition, radiometric labels suffer from the drawbacks of short useful life and handling hazards.
Labeling with magnetic iron oxide has also been proposed, as described in U.S. Pat. No. 4,452,773 (issued June 5, 1984 to Molday). The sensitivity of such labels is also limited, and their use in labeling biological species requires expensive equipment and tedious procedures.
Fluorescent spectroscopy, one of the most sensitive and versatile of the optical analytical techniques, has become increasingly popular in recent years to overcome drawbacks of other labeling techniques. In fluorescence spectroscopy, a sample containing a fluorescent species is irradiated with light of known spectral distribution within the exitation spectrum of the target fluorescent species. The intensity of the resulting characteristic emission spectrum of the fluorescent target molecules is determined and is related to the number of target molecules in the sample. Fluorescent spectroscopy is used extensively for studies of protein structure, bacterial cell wall reactions and conformational changes in enzymes, as well as for determinations of an immunologically reactive ligand in a specific binding assay.
Fluorescent labels comprising chelates of a rare earth element incorporated into polymeric particles of a loadable latex are described in U.S. Pat. Nos. 4,259,313 (issued Mar. 31, 1981 to Frank et al) and related 4,283,382 (issued Aug. 11, 1981 to Frank et al). These labels exhibit improved efficiency in fluorescence and are particularly useful for immunoassays. The polymeric particles serve as carriers for immunologically reactive species directly attached thereto.
Our colleagues, J. R. Schaeffer, T. J. Chen and M. A. Schen, have discovered that certain polymers provide exceptionally stable fluorescent labels. These labels are the subject of copending and commonly assigned U.S. Ser. No. 713,202 filed Mar. 18, 1985 and entitled STABILIZED FLUORESCENT RARE EARTH LABELS AND LABELED PHYSIOLOGICALLY REACTIVE SPECIES. However, these polymers comprise a small class of materials composed of recurring units derived from certain combination of monomers.
Although the labels of the Frank et al references represent a breakthrough in clinical chemistry because of their improved fluorescence efficiency, there is a need to render the broad class of polymeric labels described therein more stable in aqueous solutions. The labels of Frank et al generally tend to agglutinate spontaneously and to settle out of solution. They therefore have a shortened storage life. They also demonstrate a tendency to agglutinate prematurely during an assay. The improvements discovered by Schaeffer et al are limited to a small class of polymeric labels. Hence, it is desirable to have fluorescent labels which avoid the above problems.